Information on the mechanisms of cell differentiation which lead to mineral deposition is essential for an understanding of both normal craniofacial development and diseases associated with bone formation and removal. Mineralizing tissues undergo marked cellular changes during matrix mineral- ization. Among the observed changes are altered cell morphology, metaboli- sm and gene expression. Many components have been implicated in inducing cells to undergo these mineralization-specific changes including matrix vesicle release, change in cellular redox status and appearance of mineral- ization-specific mRNAs and proteins. Elevated alkaline phosphatase levels are a hallmark of mineralization in both bone and cartilage, and appearance of type X collagen is correlated with the onset of mineralization in cartilage. Factors leading to formation of mineralized matrix have been extensively studied in cultured osteoblasts and, to a lesser extent, in cultured chondrocytes. Wit both cell types, ascorbic acid can induce elevated alkaline phosphatase as well as mineral deposition; this mineralization is potentiated in the presence of the organic phosphate donor beta-glyceropho- sphate. We have shown that in cultured chondrocytes, ascorbic acid will also induce a transition from type II collagen to type X collagen. One effect of ascorbate is its well-documented influence on procollagen hydroxylation and secretion. However, the vitamin may also affect gene transcription as well as cellular metabolism. The primary goal of this research is to further our understanding of events controlling formation of calcified tissues by defining the factors in- fluencing matrix mineralization in cultured chondrocytes. These studies will examine the hypotheses that ascorbate-induced changes in mineralizing cultures are mediated by effects on matrix composition, cell-matrix interaction and/or cell metabolism. They will also utilize chondrocyte transfection assays to test the hypothesis that high levels of alkaline phosphatase or type X collagen induce changes leading to mineralization.